The effects adiposity, diabetic status and depot-specificity on the activation of NFKB on JNK in human abdominal adipose tissue

Background and Aims: Both serum adiponectin, an adipokine with insulinsensitizing and anti-inflammatory action, and high sensitivity C-reactive protein (hsCRP), a marker of chronic low-grade systemic inflammation, have been reported to predict the development of type 2 diabetes. Here we examined, in a 10-year prospective study, whether these two biomarkers had an additive effect on predicting the long-term risk of type 2 diabetes. Materials and Methods: Serum adiponectin and hpCRP were measured in 417 non-diabetic Chinese subjects from the Hong Kong Cardiovascular Risk Factor Prevalence Study, who had returned for their 10-year follow-up in 2005/06. Of these, 76 had developed DM, according to the 1998 WHO diagnostic criteria. Adiponectin was measured using an in-house ELISA assay and hsCRP was measured using a particle-enhanced immunoturbidimetric assay. The association of adiponectin and hsCRP, alone or in combination, with the 10-year risk of diabetes was investigated. Results: Compared to the 341 subjects who remained non-diabetic at year- 10, subjects who had developed diabetes consisted of more men (p<0.003). They had higher baseline BMI, waist circumference, mean arterial blood pressure, plasma glucose (fasting and 2-hour post-OGTT), fasting insulin, HOMA-insulin resistance index (HOMA-IR), triglycerides, LDL-cholesterol and hsCRP levels (all p<0.001 except for LDL-cholesterol with p<0.05). They also had lower baseline serum adiponectin and HDL-cholesterol levels (both p<0.001). In a stepwise multiple logistic regression analysis model including sex, age, BMI, fasting insulin, presence of metabolic syndrome according to NCEP criteria, adiponectin and hsCRP, only baseline adiponectin (adjusted OR 0.44; 95% CI 0.27–0.74; p=0.002), hsCRP (adjusted OR 1.43; 95% CI 1.10–1.84; p=0.007) and metabolic syndrome (adjusted OR 2.72; 95% CI 1.55–4.78; p=0.001) were independently predictive of diabetes at 10 years. The addition of baseline adiponectin or hsCRP to a model consisting of sex, age and BMI significantly improved the prediction of diabetes risk, as reflected by the likelihood ratios in logistic regression analysis (p=0.001 for adiponectin and p=0.019 for hsCRP). Furthermore, the combined addition of adiponectin and hsCRP to the model provided significantly greater improvement than the addition of adiponectin alone (p=0.048) or the addition of hsCRP alone (p=0.003). Conclusion: Serum adiponectin and hsCRP were both independent predictors of the 10-year risk of diabetes in this Chinese cohort. Their usefulness as biomarkers for predicting the development of type 2 diabetes were over and above those of conventional risk factors including sex, age and BMI, and appeared to be additive when these two new biomarkers were used together. Supported by the STR seeding fund on Healthy Aging of the University of Hong Kong and the Innovation & Technology Fundlink_to_subscribed_fulltex

Long-term orlistat treatment reduces endotoxinaemia in fatty liver disease patients

Background and Aim: We previously tested the effects of a 18h culture in 2, 5, 10 and 30 mmol/l glucose (G2, G5, G10 and G30) on the transcriptome of cultured rat islets and identified 18 clusters with distinct glucose-dependent mRNA profiles. Of these, genes that were up-regulated between G10 and G30 are particularly interesting, as they may contribute to beta-cell glucotoxicity or protection against it. Besides genes involved in the unfolded protein response, this cluster contains most glycolytic enzymes and other genes typically induced by hypoxia, such as Adrenomedullin (Adm). Hypothesis: High glucose, which increases O2 consumption in β-cells, may induce hypoxia and thereby cause glucotoxicity. In this study, we tested whether overnight culture in high glucose activates the hypoxia-inducible transcription factor HIF in cultured rat beta-cells. Material and Methods: Male Wistar rat islets were precultured for one week in serum-free RPMI medium containing 10 mmol/l glucose (G10) and 5 g/l BSA, during which islets with signs of central necrosis were systematically discarded. INS1-E cells were cultured as monolayers (passage 72 to 80) in the presence of 10% FBS. Rat islets and INS1-E cells (70% confluence) were then cultured 18h in the presence of increasing glucose concentrations (G2, G5, G10 and G30) with various test substances and at different O2 concentrations (1%, 20% or 60% corresponding to a pO2 of 7.6, 152 and 456 mmHg). Results: Compared with G2, culture in G30 significantly increased 4-fold the protein levels of HIF1α and HIF2α, and 2-fold their dimerization partner HIF1β in INS1-E cell nuclear extracts while significantly decreasing HIF1β protein levels by ~50% in cytosolic extracts. High glucose also significantly up-regulated the mRNA levels of several HIF target genes (Adm, Glyceraldehyde phosphate dehydrogenase, Aldolase A, Lactate dehydrogenase A). These glucose effects, which were confirmed in whole rat islets, were mimicked by 6h exposure to a low pO2 (1% O2) or by 18h treatment with 10-30 µmol/l CoCl2, a known activator of HIF. In contrast, they were almost completely inhibited by 60% O2 or by various agents that inhibit Ca2+ influx and insulin secretion (250 µmol/l diazoxide, 1 µmol/l nimodipine and 1 µmol/l clonidine) and thereby likely reduce without suppressing the glucose stimulation of β-cell O2 consumption. In comparison with HIF target genes, Thioredoxin interacting protein (Txnip), one of the most glucose-responsive genes in cultured rat islets, was regulated in a completely different manner. Thus, Txnip mRNA levels were not increased by CoCl2 and 1% O2, and their increase by G30 was not reduced by 60% O2 and was markedly enhanced by diazoxide, nimodipine and clonidine. Conclusion: High glucose induces HIF1α and HIF2α protein stabilization, HIF1/2α-HIF1β dimer translocation to the nucleus, and increased expression of glycolytic enzymes and other HIF target genes in cultured rat beta-cells. These effects likely result from the glucose stimulation of ATP utilization and O2 consumption which may induce beta-cell hypoxia. These effects could contribute to in vitro β-cell glucotoxicity not only in whole islets but also in insulin-secreting cells cultured as monolayers

Visfatin is regulated by rosiglitazone in type 2 diabetes mellitus and influenced by NFκB and JNK in human abdominal subcutaneous adipocytes

Visfatin has been proposed as an insulin-mimicking adipocytokine, predominantly secreted from adipose tissue and correlated with obesity. However, recent studies suggest visfatin may act as a proinflammatory cytokine. Our studies sought to determine the significance of this adipocytokine and its potential role in the pathogenesis of T2DM. Firstly, we examined the effects of diabetic status on circulating visfatin levels, and several other adipocytokines, demonstrating that diabetic status increased visfatin*, TNF-α*** and IL-6*** compared with non-diabetic subjects (*p<0.05, **p<0.01, ***p<0.001, respectively). We then assessed the effects of an insulin sensitizer, rosiglitazone (RSG), in treatment naïve T2DM subjects, on circulating visfatin levels. Our findings showed that visfatin was reduced post-RSG treatment [vs. pre-treatment (*p<0.05)] accompanied by a reduction in HOMA-IR**, thus implicating a role for insulin in visfatin regulation. Further studies addressed the intracellular mechanisms by which visfatin may be regulated, and may exert pro-inflammatory effects, in human abdominal subcutaneous (Abd Sc) adipocytes. Following insulin (Ins) and RSG treatment, our in vitro findings highlighted that insulin (100 nM), alone, upregulated visfatin protein expression whereas, in combination with RSG (10 nM), it reduced visfatin*, IKKβ** and p-JNK1/2*. Furthermore, inhibition of JNK protein exacted a significant reduction in visfatin expression (**p<0.01), whilst NF-κB blockade increased visfatin (*p<0.05), thus identifying JNK as the more influential factor in visfatin regulation. Additional in vitro analysis on adipokines regulating visfatin showed that only Abd Sc adipocytes treated with recombinant human (rh)IL-6 increased visfatin protein (*p<0.05), whilst rh visfatin treatment, itself, had no influence on TNF-α, IL-6 or resistin secretion from Sc adipocytes. These data highlight visfatin's regulation by insulin and RSG, potentially acting through NF-κB and JNK mechanisms, with only rh IL-6 modestly affecting visfatin regulation. Taken together, these findings suggest that visfatin may represent a pro-inflammatory cytokine that is influenced by insulin/insulin sensitivity via the NF-κB and JNK pathways